SPOT 4.1 Basic and Advanced Software Manual for SPOT Insight ...

Ch. 5 – Preparing to Take a PictureA thermal frame is a longer exposure dark frame with the bias frame subtracted. All that is left aftersubtraction of the bias frame is the accumulation of dark current electrons in each pixel that happenedduring the exposure time of the dark frame.Subtracting a thermal frame from any image of the same exposure time will remove the dark currentelectrons that accumulated during the exposure time. Since the accumulation of dark current electrons isdirectly proportional to exposure time (twice the exposure time results in twice as many dark currentelectrons) the thermal frame is “scalable”. This means that one thermal frame (at a known exposure time)can be scaled up or down to predict the amount of dark current electrons that will accumulate in each pixelof an image of ANY exposure time.Subtracting both a bias frame and a (scaled) thermal frame should erase the system offset and eliminate allelectrons due to dark current and leave an image that has pixel values due only to light striking the CCD.This is a highly useful way to “clean up” an image.Get Bias Frame (Bias Frame Subtraction)Get Bias Frame allows you to collect and store a bias frame. Bias frame subtraction is a process that usesthe stored bias frame, subtracting it from a captured image to remove both the system offset and the darkcurrent electrons that accumulated in each pixel while it was waiting to be read out. When this subtractionprocess has been applied to a short exposure image, the remaining data (and image) is mostly representativeof the true image data being captured.You may select a bias frame to use in the subtraction process within the image set-up post-processingwindow. Select the checkbox for Bias Frame and then browse to select a Bias Frame correction file fromthose that have been previously developed and stored.For a step-by-step explanation for acquiring bias frames, refer to Ch. 7 – Taking a Picture.Note: Bias frame subtraction is available only when a monochrome camera is set up in 12 bitcapture and black level subtract is not selected.Get Thermal Frame (Thermal Frame Subtraction)Get Thermal Frame is a function that allows you to collect data to eliminate the dark current accumulatedduring the exposure time of your captured image. By determining the rate of dark current accumulation ineach pixel, we can scale the quantitative effect of the dark current for each pixel in the image plane (theThermal Frame) based on the time of image exposure and subtract it from the final image. The end result isa processed image that more closely reflects the true data that is collected from the specimen during theimage capture.The SPOT program accomplishes this method of thermal frame characterization by collecting one, or anaverage of a series of images that reflect the electrons accumulated due to the temperature of the CCD.This information is then stored as the Thermal Frame data. For a given experimental condition (i.e.temperature in the room), normally only one set of Thermal Frame images must be collected, with its datalater scaled to correspond to the time of the exposure.Note: Collecting thermal frames should be done from time to time depending upon the changingnature of the environment that the camera is in.Because Insight cameras are not temperature regulated, the temperature of the CCD may vary overtime. This variation will also change the dark current characteristics of the CCD. Because thermalframes capture the dark current rate at a certain CCD temperature, thermal frames should becollected as close to the image capture period as possible.User Guide to the SPOT Insight Camera 111